BACKGROUND AND AIMS: Inositol 1,4,5-trisphosphate receptor type 1 (IPR1) has been proposed to play a physiological role in regulating gastrointestinal (GI) motility, but the underlying cell-dependent mechanism remains unclear. Here, we utilized cell-specific IPR1 deletion strategies to address this question in mice. METHODS: Conditional IPR1 knockout mice using Wnt1-Cre, Islet1-Cre mice, and smMHC-Cre were generated. Cell lineage tracing was performed to determine where gene deletion occurred in the GI tract. Whole-gut transit assay and isometric tension recording were used to assess GI function in vivo and in vitro. RESULTS: In the mouse GI tract, Islet1-Cre targeted smooth muscle cells (SMCs) and interstitial cells of Cajal (ICCs), but not enteric neurons. IPR1 deletion by Islet1-Cre (isR1KO) caused a phenotype of intestinal pseudo-obstruction (IPO), evidenced by prolonged whole-gut transit time, enlarged GI tract, abdominal distention, and early lethality. IPR1 deletion by Islet1-Cre not only reduced the frequency of spontaneous contractions but also decreased the contractile responses to the muscarinic agonist carbachol (CCh) and electrical field stimulation (EFS) in colonic circular muscles. By contrast, smMHC-Cre only targeted SMCs in the mouse GI tract. Although IPR1 deletion by smMHC-Cre (smR1KO) also reduced the contractile responses to CCh and EFS in colonic circular muscles, the frequency of spontaneous contractions was less affected, and neither global GI abnormalities nor early lethality was found in smR1KO mice. CONCLUSIONS: IPR1 deletion in both ICCs and SMCs but not in SMCs alone causes an IPO phenotype, suggesting that IPR1 in ICCs plays an essential role in regulating GI motility in vivo.